Figure 1. Ribbon representation of the hemagglutinin HA0 trimer from the 1918
influenza virus. Each monomer possesses two important sites: 1) the 'Receptor
binding site' (blue shade) for virus attachment to the host lung epithelial
cells via sialic acid containing host cell receptors. 2) the 'Cleavage site'
where for full infectivity, the single chain (HA0) is cut into two chains (HA1
colored red and HA2 colored green). At the N-terminal end of the HA2 chain is
the fusion peptide which is critical for subsequent membrane fusion events that
lead to infection. The 1918 HA monomer also possesses 2 basic patches (orange
shade) which may have contributed to the increased infectivity observed during
the pandemic.

In 1918, a Great Flu Pandemic swept the world, killing an estimated 20-40
million people, thus making it the largest and most destructive outbreak of any
infectious disease in recorded history. This virus was unusual not only in its
higher than average mortality rate, but also in the age groups that it
attacked. Unlike other outbreaks it targeted not only the very young and old,
but also young adults (20-40 year olds). Why the 1918 virus was so devastating
is still a mystery. A thorough molecular understanding is now sought to offer
reasons why this influenza virus was so pathogenic and how it managed to evade
the immune system so effectively. Unfortunately, no intact virus survived,
since the pandemic struck before viruses were even identified as the causative
agent. Only fragments of the viral genome survived in both Alaskan victims
buried in the permafrost and in fixed and archived autopsy material and these
have been used to sequence and assemble a number of genes from this virus.

One of the most important viral proteins is the virus coat protein,
hemagglutinin (HA). This surface glycoprotein is responsible for virus binding
to host cell receptors, and subsequent membrane fusion events within the
endosomal pathway in the infected cell. HA is also the most abundant antigen on
the surface of the virus and harbors the primary neutralizing epitopes for
antibodies. In line with its importance, this viral antigen has been the
initial focus of our research on the 1918 flu.

The ectodomain of the HA gene from the 1918 influenza virus1 A/South Carolina/1/18 was cloned and expressed in a
baculovirus expression system as the pre-infective HA0 form (see Figure 1
legend). Protein was produced as trimers using a trimerizing sequence
('foldon') from the bacteriophage T4 fibritin. HA0 crystallized at pH 5.5 and
its structure was recently solved by molecular replacement (MR) to 3.0 ┼
resolution from data collected at beamline 9-2 at SSRL (pdb:1RD8)2. 18HA0 is ~135 ┼ in length with two distinct domains
(Figure 1). The cylindrical trimer has a tightly intertwined 'stem' domain at
its membrane proximal base. The dominant feature of this stalk region is a long
triple-stranded coiled-coil. This region also contains the cleavage site where
host enzymes cleave HA0 to its infective HA1/HA2 form. The membrane-distal
domain consists of a globular 'head' which contains the host receptor binding
site and major epitopes for neutralizing antibodies.

Although phylogenetic analyses place the 1918 HA sequence at the base of the
evolutionary tree of human viruses, analysis of this structure revealed that it
is more closely akin to avian forms. A number of species- and serotype-specific
features have been identified:
1.
A narrow, avian-like receptor-binding site predominates in the 1918 HA. Within
this pocket, the only difference between 1918 HA and known swine-avian adapted
H1 viruses is an Glu190Asp mutation, which leads to a minimal
increase in the pocket size that could perhaps increase affinity for human
receptors.
2.
Two previously unobserved histidine-rich basic patches may enhance the
pH-dependent viral-fusion event within the endosomal pathway, required for
influenza infection. One patch, unique to both human and avian H1, H2 and H5
subtypes, is adjacent to the cleavage site, and may be involved in either
trimer destabilization or expulsion of the fusion peptide prior to the membrane
fusion event. The second patch situated at the base of the HA1 globular domain
is found only in avian H1 subtypes, providing tantalizing evidence of a direct
jump of this virus from birds to the human na´ve population.

Thus, these and other as yet unidentified features may have contributed to the
extraordinarily high infectivity and mortality rates observed during 1918.